I combine optogenetic stimulation with multi-optetrode recordings in freely moving mice to study how reward associative learning and neural activities in the ventral tegmental area (VTA) are modulated by VTA afferents. The VTA plays an essential role in reward, motivation and associative learning. The mesolimbic dopamine (DA) system helps establish incentive value for neutral stimulus in acquisition of stimulus-reward associations. Dopamine neurons fire phasic burst activity during presentation of reward or reward predictive cues. It is believed that DA neuron burst activity depends on afferent inputs. Anatomical and physiological evidence lead me to study the regulatory roles of the pedunculopontine tegmental nucleus (PPTg) in VTA circuits and its role in modulating behaviors. I employed optogenetic approach to selectively inhibit PPTg glutamatergic or cholinergic inputs to the VTA in order to examine the contribution of these inputs in Pavlovian appetitive conditioning. Furthermore, I performed in vivo optetrode recording in freely moving mice as they performed a Pavlovian task to examine the regulation of VTA circuits by PPTg glutamatergic or cholinergic afferents. The results show that PPTg-to-VTA excitatory inputs play a critical role in the acquisition of stimulus-reward associations by dampening cue discrimination. Surprisingly, the results from in vivo recording experiments show that cue-elicited burst activity of DA neurons is not affected by photoinhibition of PPTg-to-VTA excitatory inputs. Instead, I observe that photoinhibition significantly impacts cue-elicited neural activity in non-DA neurons. This raised a possibility that VTA non-DA neurons may play a role in acquisition of cue-reward association. To examine this possibility, VTA GABAergic or glutamatergic neural activity is inhibited with optogenetic inhibition approach during Pavlovian conditioning learning. The results show that inhibition of VTA GABAergic or glutamatergic neural activity dampens cue-reward associative learning. In conclusion, my study reveals an unprecedented role of PPTg-to-VTA pathways in modulating the activity of VTA non-DA neurons and cue-reward associative learning. My future research will build on these findings to explore possible involvement of VTA non-DA efferents in cue-reward associative learning. The goals are to determine which putative targets innervated by VTA non-DA neurons are critically required in discrimination learning, whether chronic use of substance of abuse will dampen discrimination learning and impact selective VTA non-DA efferents and develop an optical intervention technique to rescue learning deficits.